Virus polypeptide-protein subunit combination vaccine based on dna nanotechnology, and preparation method therefor and use thereof

Abstract

Provided are a virus peptide-protein subunit combination vaccine based on DNA nanotechnology, and a preparation method therefor and use thereof, which belong to the technical field of biological products. Provided is a virus peptide-protein subunit combination vaccine based on DNA nanotechnology, in which a tetrahedral framework nucleic acid formed by assembly of DNA strands is used as a vector, a protein antigen triggering virus-specific T cell activation is coupled to one edge of the tetrahedral framework nucleic acid, and B cell epitope peptides of a virus are coupled to four vertices of the tetrahedral framework nucleic acid.

1 . A virus peptide-protein subunit combination vaccine based on DNA nanotechnology, wherein a tetrahedral framework nucleic acid formed by assembly of DNA strands is used as a vector, a protein antigen triggering virus-specific T cell activation is coupled to one edge of the tetrahedral framework nucleic acid, and B cell epitope peptides of a virus are coupled to four vertices of the tetrahedral framework nucleic acid. 
     
     
         2 . The virus peptide-protein subunit combination vaccine based on DNA nanotechnology according to  claim 1 , wherein nucleotide sequences of the DNA strands for assembly of a tetrahedral framework nucleic acid are set forth in SEQ ID NO: 1 to SEQ ID NO: 8. 
     
     
         3 . The virus peptide-protein subunit combination vaccine based on DNA nanotechnology according to  claim 1 , wherein the B cell epitope peptides of a virus are hybridized, via a sulfhydryl-modified DNA strand, with a single-stranded DNA protruding from the vertices of the tetrahedral framework nucleic acid;
 the nucleotide sequence of the sulfhydryl-modified DNA strand is set forth in SEQ ID NO: 9.   
     
     
         4 . The virus peptide-protein subunit combination vaccine based on DNA nanotechnology according to  claim 1 , wherein the protein antigen triggering virus-specific T cell activation is hybridized, via a DNA strand coupled with it, with a single-stranded DNA protruding from the edge of the tetrahedral framework nucleic acid;
 the nucleotide sequence of the coupled DNA strand is set forth in SEQ ID NO: 10.   
     
     
         5 . The virus peptide-protein subunit combination vaccine based on DNA nanotechnology according to  claim 4 , wherein the protein antigen triggering virus-specific T cell activation and the DNA strand are coupled via a Halo tag and a Halo ligand. 
     
     
         6 . The virus peptide-protein subunit combination vaccine based on DNA nanotechnology according to  claim 1 , wherein the virus comprises at least one of the following viruses: a coronavirus, a human immunodeficiency virus peptide vaccine, a respiratory syncytial virus peptide vaccine and an influenza virus. 
     
     
         7 . The virus peptide-protein subunit combination vaccine based on DNA nanotechnology according to  claim 6 , wherein the coronavirus comprises a novel coronavirus and a variant thereof;
 a protein antigen triggering virus-specific T cell activation in the novel coronavirus or the variant thereof comprises an N protein;   a B cell epitope peptide of the novel coronavirus or the variant thereof comprises S3 404-412  and/or S4 440-445 .   
     
     
         8 . A method for preparing the virus peptide-protein subunit combination vaccine based on DNA nanotechnology according to  claim 1 , comprising the following steps:
 performing a first hybridization reaction on DNA strands for assembly of a tetrahedral framework nucleic acid to obtain a tetrahedral framework nucleic acid;   mixing a sulfhydryl-modified DNA strand with a B cell epitope peptide of a maleimide-modified virus and performing a coupling reaction to obtain a DNA-coupled B cell epitope peptide;   subjecting a Halo tag-modified protein antigen triggering virus-specific T cell activation to covalent coupling with a Halo ligand-modified DNA strand to obtain a DNA-coupled protein antigen;   subjecting the tetrahedral framework nucleic acid, the DNA-coupled B cell epitope peptide and the DNA-coupled protein antigen to a second hybridization reaction to obtain the virus peptide-protein subunit combination vaccine.   
     
     
         9 . The preparation method according to  claim 8 , wherein in the first hybridization reaction, the DNA strands for assembly of a tetrahedral framework nucleic acid are mixed at an equimolar ratio, treated at 95° C. for 10 min, and then maintained at 4° C. for 30 min; a final concentration of each of the DNA strands for assembly of a tetrahedral framework nucleic acid is independently 0.8-1.2 μM;
 in the coupling reaction, a molar ratio of the sulfhydryl-modified DNA strand to the maleimide-modified B cell epitope peptide of a virus is 1:(2-10); 
 in the covalent coupling, a molar ratio of the Halo tag-modified protein antigen triggering virus-specific T cell activation to the Halo ligand-modified DNA strand is (0.9-1.1):(0.9-1.1); 
 in the second hybridization reaction, a molar ratio of the tetrahedral framework nucleic acid, the DNA-coupled B cell epitope peptide and the DNA-coupled protein antigen is (0.9-1.1):(3.9-4.1):(0.9-1.1). 
 
     
     
         10 . (canceled) 
     
     
         11 . The method according to  claim 8 , wherein nucleotide sequences of the DNA strands for assembly of a tetrahedral framework nucleic acid are set forth in SEQ ID NO: 1 to SEQ ID NO: 8. 
     
     
         12 . The method according to  claim 8 , wherein the B cell epitope peptides of a virus are hybridized, via a sulfhydryl-modified DNA strand, with a single-stranded DNA protruding from the vertices of the tetrahedral framework nucleic acid;
 the nucleotide sequences of the sulfhydryl-modified DNA strands are set forth in SEQ ID NO: 9.   
     
     
         13 . The method according to  claim 8 , wherein the protein antigen triggering virus-specific T cell activation is hybridized, via a DNA strand coupled with it, with a single-stranded DNA protruding from the edge of the tetrahedral framework nucleic acid;
 the nucleotide sequence of the coupled DNA strand is set forth in SEQ ID NO: 10.   
     
     
         14 . The method according to  claim 13 , wherein the protein antigen triggering virus-specific T cell activation and the DNA strand are coupled via a Halo tag and a Halo ligand. 
     
     
         15 . The method according to  claim 8 , wherein the virus comprises at least one of the following viruses: a coronavirus, a human immunodeficiency virus peptide vaccine, a respiratory syncytial virus peptide vaccine and an influenza virus. 
     
     
         16 . The method according to  claim 15 , wherein the coronavirus comprises a novel coronavirus and a variant thereof;
 a protein antigen triggering virus-specific T cell activation in the novel coronavirus or the variant thereof comprises an N protein;   a B cell epitope peptide of the novel coronavirus or the variant thereof comprises S3 404-412  and/or S4 440-445 .